Biosensor-integrated transposon mutagenesis reveals <i>rv0158</i> as a coordinator of redox homeostasis in <i>Mycobacterium tuberculosis</i>.

Abstract

<i>Mycobacterium tuberculosis</i> (<i>Mtb</i>) is evolutionarily equipped to resist exogenous reactive oxygen species (ROS) but shows vulnerability to an increase in endogenous ROS (eROS). Since eROS is an unavoidable consequence of aerobic metabolism, understanding how <i>Mtb</i> manages eROS levels is essential yet needs to be characterized. By combining the Mrx1-roGFP2 redox biosensor with transposon mutagenesis, we identified 368 genes (redoxosome) responsible for maintaining homeostatic levels of eROS in <i>Mtb</i>. Integrating redoxosome with a global network of transcriptional regulators revealed a hypothetical protein (Rv0158) as a critical node managing eROS in <i>Mtb</i>. Disruption of <i>rv0158</i> (<i>rv0158</i> KO) impaired growth, redox balance, respiration, and metabolism of <i>Mtb</i> on glucose but not on fatty acids. Importantly, <i>rv0158</i> KO exhibited enhanced growth on propionate, and the Rv0158 protein directly binds to methylmalonyl-CoA, a key intermediate in propionate catabolism. Metabolite profiling, ChIP-Seq, and gene-expression analyses indicate that Rv0158 manages metabolic neutralization of propionate toxicity by regulating the methylcitrate cycle. Disruption of <i>rv0158</i> enhanced the sensitivity of <i>Mtb</i> to oxidative stress, nitric oxide, and anti-TB drugs. Lastly, <i>rv0158</i> KO showed poor survival in macrophages and persistence defect in mice. Our results suggest that Rv0158 is a metabolic integrator for carbon metabolism and redox balance in <i>Mtb</i>.